Project Summary Rotavirus is the leading cause of severe childhood gastroenteritis, resulting in over 215,000 deaths per year. The recently introduced live, oral rotavirus vaccine (RVV) demonstrated 90-95% protection in Europe and the US, but is less than 50% in low-income countries, where the disease burden is highest. The biological basis of rotavirus vaccine failure remains unknown but likely includes both microbial and host factors. In Malawi there is a high prevalence of rotavirus disease and after a pivotal placebo-controlled clinical trial there was modest seroconversion, only 40%-50%. Despite high vaccine coverage rotavirus remains the leading cause of diarrhea hospitalization among infants in Malawi. There is no evidence of vaccine escape however potential explanations for RVV failure include: effects of gut/systemic infection, variation in the microbiome, interference from passively acquired maternal antibodies, and/or genetic predisposition but evidence for each of these is limited. Preliminary data suggest that mutations in blood group antigens may influence rotavirus vaccine failure, but this hypothesis has not been well characterized in African populations. There is additional evidence that the high pathogen burden in low-income countries may be reducing infants' response to live vaccines. To fully understand the causes of RVV failure a robust investigation of host and microbial genetic factors is needed. To elucidate genetic variants linked to low rotavirus vaccine immunogenicity and clinical vaccine failure, this study will utilize a genome wide association study (GWAS) and transcriptomic analysis. Detailed infant health will be collected at baseline and established diagnostic assays will be used to characterize serological response to RVV. To assess whether concurrent infection at the time of vaccination is correlated with RVV failure, common pathogens will be screened for ? using commercially available diagnostics, novel CRISPR- Cas13a assays, and metagenomic sequencing . This study will leverage prospective and retrospective collections combined with a GWAS and transcriptomic profiling of infants pre vs. post-vaccination to identify genetic variants that are associated with and predictive of RVV failure. The genetic variants identified in the GWAS will undergo improved signal resolution and functional analysis through Composite of Multiple Signals, Massively Parallel Report Assay, In vitro analysis, and gene editing using CRISPR. This study will produce the first GWAS of a viral diarrheal illness in Africa and may uncover mechanisms of defense to rotavirus. Despite widespread vaccination, rotavirus remains the main cause of severe life threatening diarrhea in Malawian infants. This study has the potential to identify host-derived signatures that can be used as early predictors of RVV failure, inform immunization strategies and the future of vaccine development.